#METABOLOMICS WORKBENCH TatianaJoao_20220509_094827 DATATRACK_ID:3234 STUDY_ID:ST002202 ANALYSIS_ID:AN003605 PROJECT_ID:PR001405 VERSION 1 CREATED_ON June 29, 2022, 6:30 am #PROJECT PR:PROJECT_TITLE Metabolic profiling of induced acute pancreatitis and pancreatic cancer PR:PROJECT_TITLE progression in a mutant Kras mouse model PR:PROJECT_TYPE NMR-based metabolomics PR:PROJECT_SUMMARY Untargeted Nuclear Magnetic Resonance (NMR) metabolomics of polar extracts from PR:PROJECT_SUMMARY the pancreata of a caerulin-induced mouse model of pancreatitis (Pt) and of a PR:PROJECT_SUMMARY transgenic mouse model of pancreatic cancer (PCa) were used to find metabolic PR:PROJECT_SUMMARY markers of Pt and to characterize the metabolic changes accompanying PCa PR:PROJECT_SUMMARY progression. Using multivariate analysis a 10-metabolite metabolic signature PR:PROJECT_SUMMARY specific to Pt tissue was found to distinguish the benign condition from both PR:PROJECT_SUMMARY normal tissue and precancerous tissue (low grade pancreatic intraepithelial PR:PROJECT_SUMMARY neoplasia, PanIN, lesions). PR:INSTITUTE University of Aveiro PR:DEPARTMENT Department of Chemistry and CICECO-Aveiro Institute of Materials PR:LABORATORY Metabolomics from Ana M. Gil PR:LAST_NAME Carneiro PR:FIRST_NAME Tatiana J. PR:ADDRESS Campus Universitário de Santiago, Aveiro, Aveiro, 3810-193, Portugal PR:EMAIL tatiana.joao@ua.pt PR:PHONE +351 234 370 200 #STUDY ST:STUDY_TITLE Metabolic profiling of induced acute pancreatitis and pancreatic cancer ST:STUDY_TITLE progression in a mutant Kras mouse model ST:STUDY_TYPE NMR-based metabolomics ST:STUDY_SUMMARY Untargeted Nuclear Magnetic Resonance (NMR) metabolomics of polar extracts from ST:STUDY_SUMMARY the pancreata of a caerulin-induced mouse model of pancreatitis (Pt) and of a ST:STUDY_SUMMARY transgenic mouse model of pancreatic cancer (PCa) were used to find metabolic ST:STUDY_SUMMARY markers of Pt and to characterize the metabolic changes accompanying PCa ST:STUDY_SUMMARY progression. Using multivariate analysis a 10-metabolite metabolic signature ST:STUDY_SUMMARY specific to Pt tissue was found to distinguish the benign condition from both ST:STUDY_SUMMARY normal tissue and precancerous tissue (low grade pancreatic intraepithelial ST:STUDY_SUMMARY neoplasia, PanIN, lesions). The mice pancreata showed significant changes in the ST:STUDY_SUMMARY progression from normal tissue, through low-grade and high-grade PanIN lesions ST:STUDY_SUMMARY to pancreatic ductal adenocarcinoma (PDA). These included increased lactate ST:STUDY_SUMMARY production, amino acid changes consistent with enhanced anaplerosis, decreased ST:STUDY_SUMMARY concentrations of intermediates in membrane biosynthesis (phosphocholine and ST:STUDY_SUMMARY phosphoethanolamine) and decreased glycosylated uridine phosphates, reflecting ST:STUDY_SUMMARY activation of the hexosamine biosynthesis pathway and protein glycosylation. ST:INSTITUTE University of Aveiro ST:DEPARTMENT Department of Chemistry and CICECO-Aveiro Institute of Materials ST:LABORATORY Metabolomics from Ana M. Gil ST:LAST_NAME Carneiro ST:FIRST_NAME Tatiana ST:ADDRESS Campus Universitário de Santiago, Aveiro, Aveiro, 3810-193, Portugal ST:EMAIL tatiana.joao@ua.pt ST:PHONE +351 234 370 200 ST:NUM_GROUPS 6 ST:TOTAL_SUBJECTS 40 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENOTYPE_STRAIN C57BL/6 (WT); LSL-KrasG12D/+ - p48Cre/+; LSL-KrasG12/D+; LSL-Tpr53R172H/+; SU:GENOTYPE_STRAIN Pdx-1-Cre SU:AGE_OR_AGE_RANGE 1.5 to 9 months SU:ANIMAL_LIGHT_CYCLE 12h light/dark cycles (7.00 AM lights on) SU:ANIMAL_FEED ad libitum SU:ANIMAL_WATER ad libitum SU:ANIMAL_INCLUSION_CRITERIA Healthy animails SU:SPECIES_GROUP C57BL/6 (WT); LSL-KrasG12D/+ - p48Cre/+; LSL-KrasG12/D+; LSL-Tpr53R172H/+; SU:SPECIES_GROUP Pdx-1-Cre #FACTORS #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data SUBJECT_SAMPLE_FACTORS - ES250414-N1 Treatment_group:Control RAW_FILE_NAME=ES250414-N1 SUBJECT_SAMPLE_FACTORS - ES250414-N2 Treatment_group:Control RAW_FILE_NAME=ES250414-N2 SUBJECT_SAMPLE_FACTORS - ES250414-N3 Treatment_group:Control RAW_FILE_NAME=ES250414-N3 SUBJECT_SAMPLE_FACTORS - ES250414-N4 Treatment_group:Control RAW_FILE_NAME=ES250414-N4 SUBJECT_SAMPLE_FACTORS - MB-ES-121016-22_N Treatment_group:Control RAW_FILE_NAME=MB-ES-121016-22_N SUBJECT_SAMPLE_FACTORS - MB-ES-121016-24_N Treatment_group:Control RAW_FILE_NAME=MB-ES-121016-24_N SUBJECT_SAMPLE_FACTORS - MB-ES-121016-40_N Treatment_group:Control RAW_FILE_NAME=MB-ES-121016-40_N SUBJECT_SAMPLE_FACTORS - 5848_HG Treatment_group:High-grade PanIN lesion RAW_FILE_NAME=5848_HG SUBJECT_SAMPLE_FACTORS - ES250414-HG1 Treatment_group:High-grade PanIN lesion RAW_FILE_NAME=ES250414-HG1 SUBJECT_SAMPLE_FACTORS - ES250414-HG10 Treatment_group:High-grade PanIN lesion RAW_FILE_NAME=ES250414-HG10 SUBJECT_SAMPLE_FACTORS - ES250414-HG4 Treatment_group:High-grade PanIN lesion RAW_FILE_NAME=ES250414-HG4 SUBJECT_SAMPLE_FACTORS - ES250414-HG5 Treatment_group:High-grade PanIN lesion RAW_FILE_NAME=ES250414-HG5 SUBJECT_SAMPLE_FACTORS - HG14 Treatment_group:High-grade PanIN lesion RAW_FILE_NAME=HG14 SUBJECT_SAMPLE_FACTORS - Pancreas46-HG Treatment_group:High-grade PanIN lesion RAW_FILE_NAME=Pancreas46-HG SUBJECT_SAMPLE_FACTORS - LG11 Treatment_group:Low-grade PanIN lesion RAW_FILE_NAME=LG11 SUBJECT_SAMPLE_FACTORS - LG14 Treatment_group:Low-grade PanIN lesion RAW_FILE_NAME=LG14 SUBJECT_SAMPLE_FACTORS - LG15 Treatment_group:Low-grade PanIN lesion RAW_FILE_NAME=LG15 SUBJECT_SAMPLE_FACTORS - ajw_eva5134_060616_LG Treatment_group:Low-grade PanIN lesion RAW_FILE_NAME=ajw_eva5134_060616_LG SUBJECT_SAMPLE_FACTORS - ajw_eva5846_060614_LG Treatment_group:Low-grade PanIN lesion RAW_FILE_NAME=ajw_eva5846_060614_LG SUBJECT_SAMPLE_FACTORS - ES250414-PDA-KPC Treatment_group:Pancreatic ductal adenocarcinoma RAW_FILE_NAME=ES250414-PDA-KPC SUBJECT_SAMPLE_FACTORS - ES250414-PDA1 Treatment_group:Pancreatic ductal adenocarcinoma RAW_FILE_NAME=ES250414-PDA1 SUBJECT_SAMPLE_FACTORS - MB-ES-121016-48_PDA Treatment_group:Pancreatic ductal adenocarcinoma RAW_FILE_NAME=MB-ES-121016-48_PDA SUBJECT_SAMPLE_FACTORS - MB-ES-121102-31_PDA Treatment_group:Pancreatic ductal adenocarcinoma RAW_FILE_NAME=MB-ES-121102-31_PDA SUBJECT_SAMPLE_FACTORS - Pancreas57_PDA Treatment_group:Pancreatic ductal adenocarcinoma RAW_FILE_NAME=Pancreas57_PDA SUBJECT_SAMPLE_FACTORS - Pancreas58_PDA Treatment_group:Pancreatic ductal adenocarcinoma RAW_FILE_NAME=Pancreas58_PDA SUBJECT_SAMPLE_FACTORS - Pt2 Treatment_group:Pancreatitis RAW_FILE_NAME=Pt2 SUBJECT_SAMPLE_FACTORS - Pt3 Treatment_group:Pancreatitis RAW_FILE_NAME=Pt3 SUBJECT_SAMPLE_FACTORS - Pt4 Treatment_group:Pancreatitis RAW_FILE_NAME=Pt4 SUBJECT_SAMPLE_FACTORS - Pt5 Treatment_group:Pancreatitis RAW_FILE_NAME=Pt5 SUBJECT_SAMPLE_FACTORS - Pt6 Treatment_group:Pancreatitis RAW_FILE_NAME=Pt6 SUBJECT_SAMPLE_FACTORS - Pt7 Treatment_group:Pancreatitis RAW_FILE_NAME=Pt7 SUBJECT_SAMPLE_FACTORS - ES250414-S2-sarc Treatment_group:Sarcomatoid RAW_FILE_NAME=ES250414-S2-sarc SUBJECT_SAMPLE_FACTORS - ES250414-S3-sarc Treatment_group:Sarcomatoid RAW_FILE_NAME=ES250414-S3-sarc SUBJECT_SAMPLE_FACTORS - ES250414-S4-sarc Treatment_group:Sarcomatoid RAW_FILE_NAME=ES250414-S4-sarc SUBJECT_SAMPLE_FACTORS - MB-ES-121016-30_SARC Treatment_group:Sarcomatoid RAW_FILE_NAME=MB-ES-121016-30_SARC SUBJECT_SAMPLE_FACTORS - MB-ES-121016-50_SARC Treatment_group:Sarcomatoid RAW_FILE_NAME=MB-ES-121016-50_SARC SUBJECT_SAMPLE_FACTORS - MB-ES-121016-53_SARC Treatment_group:Sarcomatoid RAW_FILE_NAME=MB-ES-121016-53_SARC SUBJECT_SAMPLE_FACTORS - MB-ES-121102-34_SARC Treatment_group:Sarcomatoid RAW_FILE_NAME=MB-ES-121102-34_SARC SUBJECT_SAMPLE_FACTORS - MB-ES-121102-37_SARC Treatment_group:Sarcomatoid RAW_FILE_NAME=MB-ES-121102-37_SARC SUBJECT_SAMPLE_FACTORS - MB-ES-121102-38_SARC Treatment_group:Sarcomatoid RAW_FILE_NAME=MB-ES-121102-38_SARC #COLLECTION CO:COLLECTION_SUMMARY Mice were sacrificed by cervical dislocation and pancreatic tissue rapidly CO:COLLECTION_SUMMARY excised and freeze-clamped using liquid nitrogen-cooled tongs. CO:SAMPLE_TYPE Pancreas #TREATMENT TR:TREATMENT_SUMMARY The control group (N, normal tissue, n=7) comprised animals that did not develop TR:TREATMENT_SUMMARY lesions or tumors (C57BL/6 wt mice); the pancreatitis group (Pt, n=6) comprised TR:TREATMENT_SUMMARY C57BL/6 wt mice in which acute pancreatitis was induced by six hourly TR:TREATMENT_SUMMARY intraperitoneal 50 ug/kg caerulein (Sigma-Aldrich, Dorset, UK) injections; the TR:TREATMENT_SUMMARY low grade lesion group (LG, n=5) comprised 4-month old KC (LSL-KrasG12D/+ - TR:TREATMENT_SUMMARY p48Cre/+) mice with predominant LG PanIN lesions (mPanIN 1 and 1A); the high TR:TREATMENT_SUMMARY grade lesion group (HG, n=7) comprised 9-month old KC mice with predominant HG TR:TREATMENT_SUMMARY PanIN lesions (mPanIN 2 and 3); the spontaneous pancreatic ductal adenocarcinoma TR:TREATMENT_SUMMARY (PDA, n=6) and sarcomatoid (Sarc., n=9) groups corresponded to 3-6 month old KPC TR:TREATMENT_SUMMARY (LSL-KrasG12/D+;LSL-Tpr53R172H/+;Pdx-1-Cre) mice. Tumors were studied when they TR:TREATMENT_SUMMARY were less than 5 mm in diameter. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Tissues (weights in the 60-120 mg range) were homogenized (ca. 200mg/mL) in 50 SP:SAMPLEPREP_SUMMARY mM HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid), 1 mM EDTA SP:SAMPLEPREP_SUMMARY (ethylenediaminetetraacetic acid), 0.7% sodium deoxy- cholate, 1% Nonidet P-40, SP:SAMPLEPREP_SUMMARY 0.5 M lithium chloride, pH 7.6, using a Precellys 24 homogeniser (Stretton SP:SAMPLEPREP_SUMMARY Scientific, Stretton, UK) and then extracted in methanol:chloroform:water. SP:PROCESSING_STORAGE_CONDITIONS -80℃ SP:EXTRACTION_METHOD Biphasic method (methanol/ chloroform/ water) SP:EXTRACT_STORAGE -80℃ #ANALYSIS AN:ANALYSIS_TYPE NMR AN:LABORATORY_NAME Metabolomics Ana M. Gil AN:SOFTWARE_VERSION Topspin 3.2 #NMR NM:INSTRUMENT_NAME Bruker 600 MHz NMR spectrometer NM:INSTRUMENT_TYPE FT-NMR NM:NMR_EXPERIMENT_TYPE 1D-1H NM:NMR_COMMENTS 1st subfolder contains raw spectra; 2nd subfolder contains manually processed NM:NMR_COMMENTS spectra NM:FIELD_FREQUENCY_LOCK H2O+D2O for 2H NM:SPECTROMETER_FREQUENCY 600MHz NM:NMR_SOLVENT D2O NM:NMR_TUBE_SIZE 5mm NM:SHIMMING_METHOD Topshim NM:RECEIVER_GAIN 724.1 NM:CHEMICAL_SHIFT_REF_CPD 3-(trimethylsilyl)-2,2’,3,3’-tetradeuteropropionic acid (TSP) NM:TEMPERATURE 298K NM:NUMBER_OF_SCANS 512 NM:ACQUISITION_TIME 4.5 s NM:RELAXATION_DELAY 12.5 s NM:SPECTRAL_WIDTH 7.3 kHz NM:NUM_DATA_POINTS_ACQUIRED 32 k NM:REAL_DATA_POINTS 64 k NM:ZERO_FILLING 64 k (and multiplied by a 0.3 Hz exponential function prior to Fourier NM:ZERO_FILLING transformation) NM:BASELINE_CORRECTION_METHOD Manual NM:CHEMICAL_SHIFT_REF_STD 3-(trimethylsilyl)-2,2’,3,3’-tetradeuteropropionic acid (TSP) NM:NMR_RESULTS_FILE ST002202_AN003605_Results.txt UNITS:ppmn #END